Parasitic nematode (PN) infections remain a major threat to human health worldwide, with more than 1 billion people infected. Children, pregnant women, and the elderly are particularly susceptible to morbidity from nematode infection. Control strategies are restricted to periodic de-worming of infected individuals, which is limited by rapid re-infection rates and the development of drug resistant worm populations. There are no vaccines available for PN infections in humans. Development of new drugs and vaccines will require a better understanding of host factors that participate in the immune response against PN infection. The requirement of an obligate host and the lack of good animal models have limited investigations into mechanisms that animal hosts employ to oppose PN attacks. Here we propose to use a system consisting of three model organisms: an insect, Drosophila melanogaster; the entomopathogenic (or insect parasitic) nematode Heterorhabditis bacteriophora; and its symbiotic bacteria Photorhabdus luminescens. This system is unique because it promises to reveal not only how pathogens evolve virulence but also how two pathogens (worm and bacteria) can synergize to exploit a common host (insect). Despite the identification and characterization of the main NF-?B immune signaling pathways in Drosophila, other evolutionary conserved pathways might regulate host immune mechanisms. It was recently shown that Transforming Growth Factor-beta (TGF-?) superfamily signals modulate the Drosophila immune response to wounding and bacterial infection and we have preliminary evidence that certain TGF-? pathway signaling molecules are potentially involved in the response against Heterorhabditis nematodes. We will use this information to investigate the exact role of TGF-? signaling components in Drosophila anti-nematode and antibacterial defense mechanisms. In Aim 1, we will analyze the Drosophila tissue-specific transcriptional response of TGF-? signaling members to Heterorhabditis and Photorhabdus, and the interaction between TGF-? signaling and other innate immune pathways. In Aim 2, we propose to examine whether TGF-? molecules modulate cellular immune functions and their interaction with humoral reactions against Heterorhabditis nematodes and their Photorhabdus bacteria. In Aim 3, we plan to study the pathologic effects caused by the two pathogens, separately or together, to Drosophila flies with compromised TGF-? signaling. We expect that the results from this project will generate novel insights into the immune role of TGF-? signaling molecules and may expose a currently unknown layer of the innate immune system. Such knowledge will contribute significantly to the development of improved practices to control PN in humans.